1,721,052 research outputs found
Type V and type III collagen modulate the expression and assembly of fibronectin extracellar matrix in classic and vascular Ehlers-Danlos syndrome fibroblasts, affecting cell survival and migration
No full textExtracellular matrix (ECM) regulates cell proliferation, migration, survival and gene expression, via signal transduction pathways differentially activated by ECM ligands interacting with specific integrins. In vitro cultured skin fibroblasts derived from classic and vascular Ehlers-Danlos syndrome (cEDS, vEDS) patients show a fibronectin (FN)-ECM disarray, consequent to the altered type V and III collagen (COLLV, COLLIII) expression and deposition. The COLL-FN-ECM disassembly is associated to the reduction of the COLLs a2b1 integrin receptor, the FN a5b1 integrin receptor substitution with avb3, rescuing from anoikis through a FAK-independent and epidermal growth factor-mediated signalling, and to the impairment of in vitro migration. The role in the ECM-mediated survival and migration of COLLV and COLLIII, respectively mutated in cEDS and vEDS, was investigated in cultured skin fibroblasts by indirect immunofluorescence, Western blotting, Q-PCR and in vitro wounding assay. Purified COLLV and COLLIII restore in cEDS and vEDS cells respectively, the COLL-ECM and the a2b1 integrin. This receptor transduces for the FN gene expression and splicing modulation, EDA+-FN isoform synthesis and organisation, a5b1-phosphorylated FAK colocalisation and EDA+-FN specific a9b1 integrin up-regulation. The induced COLL-FN-ECM, binding to a2b1-a5b1-a9b1 integrin complex, restores the FAK-mediated survival and the cell migration. These findings show the role of COLLV- and COLLIII-ECM-mediated transduction signalling in in vitro cEDS and vEDS fibroblasts’ survival and migration and give insights in the comprehension of the molecular mechanisms regulating these processes in vivo
Type III and V collagens modulate the expression and assembly of EDA+ fibronectin in the extracellular matrix of defective Ehlers-Danlos syndrome fibroblasts
No full textBackground: Alternative splicing of EDA fibronectin (FN) region is a cell type- and development-regulated
mechanism controlled by pathological processes, growth factors and extracellular matrix (ECM). Classic
and vascular Ehlers–Danlos syndrome (cEDS and vEDS) are connective tissue disorders caused by COL5A1/
COL5A2 and COL3A1 gene mutations, leading to an in vivo abnormal collagen fibrillogenesis and to an in
vitro defective organisation in the ECM of type V (COLLV) and type III collagen (COLLIII). These defects induce
the FN-ECM disarray and the decrease of COLLs and FN receptors, the α2β1 and α5β1 integrins. Purified
COLLV and COLLIII restore the COLL-FN-ECMs in both EDS cell strains.
Methods: Real-time PCR, immunofluorescence microscopy, andWestern blotting were used to investigate the
effects of COLLs on FN1 gene expression, EDA region alternative splicing, EDA+-FN-ECM assembly, α5β1
integrin and EDA+-FN-specific α9 integrin subunit organisation, α5β1 integrin and FAK co-regulation in
EDS fibroblasts.
Results: COLLV-treated cEDS and COLLIII-treated vEDS fibroblasts up-regulate the FN1 gene expression, mod-
ulate the EDA+ mRNA maturation and increase the EDA+-FN levels, thus restoring a control-like FN-ECM,
which elicits the EDA+-FN-specific α9β1 integrin organisation, recruits the α5β1 integrin and switches on
the FAK binding and phosphorylation.
Conclusion: COLLs regulate the EDA+-FN-ECM organisation at transcriptional and post-transcriptional level
and activate the α5β1–FAK complexes. COLLs also recruit the α9β1 integrin involved in the assembly of
the EDA+-FN-ECM in EDS cells.
General significance: The knowledge of the COLLs-ECM role in FN isotype expression and in EDA+-FN-ECM-
mediated signal transduction adds insights in the ECM remodelling mechanisms in EDS cells.
© 2012 Published by Elsevier B.V. 42
43
The FN13 peptide inhibits human tumor cells invasion through the modulation of alphavbeta3 integrins organization and the inactivation of ILK pathway
No full textAbstractWe report the effect of the stable expression of a 13 amino acid human fibronectin (FN) peptide (FN13) on the organization of the FN extracellular matrix (ECM) and of FN integrin receptors (FNRs), in relationship with the inhibition of cellular invasion, in three FN-ECM defective human tumor-derived cell lines: SK-Hep1C3, hepatoma, ACN, neuroblastoma, and SK-OV-3, ovary carcinoma. All these cell lines stably expressing the FN13 peptide, organized an FN-ECM, disorganized αvβ1 integrins and inactivated the ILK pathway, with the loss of secretion of MMP-9. This was associated with the inhibition of cell invasion in Matrigel matrix only in SK-Hep1C3 and ACN, but not in SK-OV-3 cells. Analysis of the integrin receptors organization showed that the FN13 expressing cells SK-Hep1C3 and ACN organized αvβ3 integrins, whereas SK-OV-3 organized αvβ5 dimers. The functional block of αvβ5 integrins, with an inactivating anti-αvβ5 antibody, led to the induction of αvβ3 integrins also in SK-OV-3 cells, and to the inhibition of cell invasion. These data show that in the human tumor cells studied FN13 inhibits the in vitro invasion through the dissociation of αvβ1 dimers, leading to ILK pathway inactivation, only when the organization of αvβ3 integrins is induced in the plasma membrane
A novel homozygous SLC2A9 mutation associated with renal-induced hypouricemia
No full textHereditary renal hypouricemia (RHUC) is a genetically heterogenous disorder characterized by defective uric acid (UA) reabsorption resulting in hypouricemia and increased fractional excretion of UA; acute kidney injury (AKI) and nephrolithiasis are recognized complications. Type 1 (RHUC1) is caused by mutations in the SLC22A12 gene, whereas RHUC2 is caused by mutations in the SLC2A9 gene. Patient ethnicity is diverse but only few Caucasian families with an SLC2A9 mutation have been reported.
The current report describes the clinical history, biochemical and molecular genetics findings of a native Austrian family with RHUC2. The propositus presented with 2 episodes of exercise-induced AKI and exhibited profound hypouricemia. Mutational screening of the SLC22A12 and SLC2A9 genes was performed.
The molecular analyses revealed the homozygous c.512G>A transition that leads to the p.Arg171His missense substitution in SLC2A9, confirming the diagnosis of RHUC2. Segregation study of the causal mutation revealed that the mother and elder sister were heterozygous carriers, whereas the younger sister was found to be homozygous.
We report the identification of a novel mutation in SLC2A9 as the cause of RHUC2 in a native Austrian family. We show that glucose transporter 9 mutations cause severe hypouricemia in homozygous individuals and confirm the high risk of AKI in male individuals harbouring these mutations. In our literature review, we provide an overview of the putative underlying pathophysiology, potential renal complications, findings on kidney biopsy as well as potential long-time renal sequela
Macrophage cell cultures for rapid isolation of intracellular bacteria: the Mycobacterium bovis model
No full textIsolation of Mycobacterium bovis from suspected cases of bovine tuberculosis demands laborious and time-consuming procedures. Also, direct PCR procedures on tissue samples show poor sensitivity, whereas radiometric and fluorescence-based identification procedures demand high running costs and do not reduce the time needed for isolation to less than 10 to 15 d. Owing to the aforementioned obstacles, the human macrophage cell line THP-1 and other macrophage cell lines were investigated in experiments of M. bovis propagation and isolation from organ samples. Macrophage cells can support a high-titered propagation within 48 h of minute amounts of both BCG and fully pathogenic M. bovis strains from organ samples. A proper antibiotic mixture prevents contamination of cell cultures. A seminested PCR for tuberculosis complex-specific insertion sequence IS6110 revealed M. bovis infection in infected cells. The same result can be obtained by a flow cytometry assay for expression of M. bovis chaperonin 10. The reduced time for isolation and identification of M. bovis (48-72 h) and the consistency of the test results make the use of macrophage cell lines attractive and cost-effective for veterinary laboratories involved in surveillance of bovine tuberculosis
L’integrina avb3 inibisce il potenziale invasivo e metastatico di cellule tumorali umane esprimenti stabilmente il peptide FN13, attraverso la disattivazione di ILK.
No full textKoolen-de Vries Syndrome: Clinical Report of an Adult and Literature Review
No full textKoolen-de Vries syndrome (KdS) is a rare genetic condition characterized by typical facial dysmorphisms, cardiac and renal defects, skeletal anomalies, developmental delay, and intellectual disability of variable level. It is caused by a 440-680-kb deletion in the 17q21.31 region, encompassing CRHR1, MAPT, IMP5, STH, and KANSL1, or by an intragenic KANSL1 mutation. The majority of the patients reported are pediatric or young adults, and long-term studies able to define the prognosis of the disease are lacking. Here, we report a patient in the fourth decade misdiagnosed in the past as classical Ehlers-Danlos syndrome for the presence of generalized joint hypermobility, who carried a 546-kb deletion in 17q21.31, and compare his phenotype with those of the few KdS adults (aged >18 years) described so far. We observed a favorable prognosis of epilepsy and cardiovascular signs and reduction of joint hypermobility with age, thus providing insight into the natural history of the disorder
Effects of IFN-alpha on the inflammatory response of swine leukocytes to bacterial endotoxin
No full textBecause low-dose interferon-alpha (IFN-alpha) treatment had proved effective in several models of chronic inflammation and autoimmune disease, a possible role of IFN-alpha in modulating the response of swine leukocytes to bacterial endotoxin was investigated in this study. Exposure of swine peripheral blood mononuclear cells (PBMC) to low concentrations of human IFN-alpha caused a strong, dose-dependent decrease in CD14 expression, the lowest level being observed at 5 U/ml IFN-alpha. This result was confirmed if PBMC were later exposed to purified lipopolysaccharide (LPS). A 10-fold lower IFN-alpha concentration (0.5 U/ml) caused the largest reduction of tumor necrosis factor-alpha (TNF-alpha) accumulation in the medium of pulmonary alveolar macrophages (PAM), stimulated with bacterial LPS. At 0.5 U/ml, the expression of the TNF-alpha gene in PAM was also strongly reduced, as opposed to cells pretreated with 50 U/ml IFN-alpha. In contrast, expression of the interleukin-1beta (IL- 1beta) gene was stimulated and that of the IL-6 gene was not significantly affected at both IFN-alpha concentrations. Results point to an important role of IFN-alpha in control of the inflammatory response to bacterial endotoxin in pigs
Collageni purificati inducono in fibroblasti umani difettivi la sintesi di fibronectina EDA+ e la sua organizzazione nella matrice extracellulare.
No full text- …
